Protoplast Fusion was Performed between Origanum Onites (Marble Thyme) and Origanum Majorana (White Thyme) Species

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Abstract:

In order to obtain a new thyme cultivars from which thyme oil can be produced more excessively, protoplast fusion was performed between Origanum onites (marble thyme) and Origanum majorana  (white thyme) species. Firstly, cells of two species were separated from their cell walls. To do this, cells were incubated overnight at dark, in CPW+mannitol and CPW+sucrose mixtures. Fusion was achieved by three times centrifugation of protoplasts (plant cells with no cell wall) in CPW+enzyme solution (20% selulase, 30% driselase, 15% pectinase). Cybrid cells were transferred from flasks to petri dishes containing M prot+enyme solution and callus cells formed after one month in growth cabinet under neutral photoperiod condition. In order to determine whether cybrid cell group is true hybrid, DNA was isolated from two thyme plants and cybrid cell group and RAPD-PCR analysis was undertaken. Eleven primers from Operon RAPD 10mer AB kit were used to reveal differences between Origanum onites and Origanum majorana plants at DNA level. Specific DNA bands belonging to two species were detected in cybrid cell group. This result proves hybridity of cybrids after protoplast fusion.

Keywords:

Plant tissue culture, Protoplast culture, RAPD-PCR analysis, Origanum onites, Origanum majorana.